Production of mannosylerythritol lipids as biosurfactants by resting cells ofCandida antarctica

Summary The resting cells ofCandida antarctica strain T-34 was found to produce a large amount of mannosylerythritol lipids as biosurfactants when incubated in the medium containing only the carbon source. The resting cells prepared from different water-soluble carbon sources were able to produce the lipids abundantly from water-insoluble carbon sources. Under the optimal conditions in a shake culture, the concentration of the total lipids amounted to about 47 g/l after 6 days, and the yield of the lipids became higher than that obtained by using the growing cells of the strain.     

Behavior of sister copies of mini-F plasmid after synchronized plasmid replication in Escherichia coli cells

To clarify whether sister copies of mini-F plasmid are immediately separated from each other after replication, we analyzed the behavior of sister mini-F copies after synchronized replication of mini-F. Sister copies of mini-F were separated immediately or shortly after replication, in contrast to sister oriC copies of the Escherichia coli chromosome.     

A case of hyperlipidemia with homozygous apolipoprotein E5 (Glu3-->Lys)

In this study, we present clinical feature of a novel case with homozygous apolipoprotein (apo) E5.The patient was a 53-year-old Japanese woman. She was from a small island off the coast of Kagoshima Prefecture, Japan. Her parents were first degree cousins. No corneal opacification, xanthomatosis, lymphadenopathy, or hepatosplenomegaly was observed. There have been no signs of clinically overt atherosclerosis to date. Her serum total cholesterol, triglycerides (TG) and high-density lipoprotein (HDL)-cholesterol levels were 11.6, 6.1 and 1.2 mmol/l, respectively, and apo A-I, A-II, B, C-II, C-III and E levels were 121, 34.8, 269, 10.4, 25.7 and 10.3 mg/dl, respectively. Serum lipoprotein profile analyzed by agarose gel electrophoresis and differential staining revealed markedly increased cholesterol and TG in both beta and prebeta-migrated lipoproteins, whereas alpha-migrated lipoprotein showed decreased cholesterol. Her apo E isoform analyzed by isoelectric focusing (IEF) was found to be homozygous apo E5.Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis of her apo E and lipoprotein lipase (LPL) genes revealed that she had a homozygous apo E (Glu3-->Lys) and heterozygous LPL variant Ser447 to Ter. Her son and daughter, both of whom had hyperlipidemia, were found to have apo E3/5 phenotype. Direct sequencing analysis of her apo E gene confirmed a homozygous one nucleotide change: G to A at nucleotide position of 2836 in the exon 3, resulting in Glu3-->Lys mutation.This is the first report of lipids and lipoprotein profiles in patients with homozygous apo E5 (Glu3-->Lys).     

Reinvestigation of the requirement of cytosolic ATP for mitochondrial protein import

Protein import into mitochondria requires the energy of ATP hydrolysis inside and/or outside mitochondria. Although the role of ATP in the mitochondrial matrix in mitochondrial protein import has been extensively studied, the role of ATP outside mitochondria (external ATP) remains only poorly characterized. Here we developed a protocol for depletion of external ATP without significantly reducing the import competence of precursor proteins synthesized in vitro with reticulocyte lysate. We tested the effects of external ATP on the import of various precursor proteins into isolated yeast mitochondria. We found that external ATP is required for maintenance of the import competence of mitochondrial precursor proteins but that, once they bind to mitochondria, the subsequent translocation of presequence-containing proteins, but not the ADP/ATP carrier, proceeds independently of external ATP. Because depletion of cytosolic Hsp70 led to a decrease in the import competence of mitochondrial precursor proteins, external ATP is likely utilized by cytosolic Hsp70. In contrast, the ADP/ATP carrier requires external ATP for efficient import into mitochondria even after binding to mitochondria, a situation that is only partly attributed to cytosolic Hsp70.     

Acceleration of matrix metalloproteinase-1 production and activation of platelet-derived growth factor receptor beta in human coronary smooth muscle cells by oxidized LDL and 4-hydroxynonenal

Increases in matrix metalloproteinases (MMPs) at atherosclerotic lesions are involved in the migration of smooth muscle cells (SMCs) into the intima and to the rupture of plaques, being implicated in the progression of atherosclerosis. The present study examined the mechanisms underlying the production of MMP-1, interstitial collagenase-1, induced by oxidized low-density lipoprotein (oxLDL) and 4-hydroxynonenal (4-HNE), factors proposed to play a pivotal role in atherogenesis, in human coronary SMCs. oxLDL promoted the production of MMP-1 with the preceding phosphorylation of extracellular signal-regulated kinase (ERK) 1/2. Immunoprecipitation of platelet-derived growth factor receptor beta (PDGFR-beta) revealed that oxLDL induced tyrosine phosphorylation of the receptor. Inhibition of the activation of PDGFR-beta and ERK1/2 resulted in a suppression of the production of MMP-1. Consistently, 4-HNE also elicited the production of MMP-1 with the preceding phosphorylation of PDGFR-beta and ERK1/2. The 4-HNE-induced production of MMP-1 was prevented when the activation of PDGFR-beta and ERK1/2 was inhibited. The present results suggest that the activation of PDGFR-beta and ERK1/2 is involved in the production of MMP-1 in oxLDL- and 4-HNE-stimulated human coronary SMCs.     

Antinociceptive effect of intrathecal administration of hypotaurine in rat models of inflammatory and neuropathic pain

Hypotaurine is an intermediate in taurine biosynthesis from cysteine in astrocytes. Although hypotaurine functions as an antioxidant and organic osmolyte, its physiological role in the central nervous system remains unclear. This study used behavioral assessments to determine whether hypotaurine influenced nociceptive transmission in acute, inflammatory, and neuropathic pain. The tail flick, paw pressure, and formalin tests were performed in male Sprague-Dawley rats to examine the effects of the intrathecal administration of hypotaurine (100, 200, 400, 600?μg) on thermal, mechanical, and chemical nociception. Chronic constriction injury (CCI) to the sciatic nerve was induced in the rats, and the electronic von Frey test and plantar test were performed to assess the effects on neuropathic pain. To determine which neurotransmitter pathway(s) was involved in the action of hypotaurine, in this study, we examined how the antagonists of spinal pain processing receptors altered the effect of 600?μg hypotaurine. To explore whether hypotaurine affected motor performance, the Rotarod test was conducted. Hypotaurine had antinociceptive effects on thermal, mechanical, and chemical nociception in the spinal cord. In CCI rats, hypotaurine alleviated mechanical allodynia and thermal hyperalgesia. These effects were reversed completely by pretreatment with an intrathecal injection of strychnine, a glycine receptor antagonist. Conversely, hypotaurine did not affect motor performance. This study demonstrated that intrathecal hypotaurine suppressed acute, inflammatory, and neuropathic pain. Hypotaurine may regulate nociceptive transmission physiologically by activating glycinergic neurons in the spinal cord, and it is a promising candidate for treating various pain states.     

Detection of mutagenic activity in particulate air pollutants.

Ames's strains of Salmonella typhimurium were used to evaluate the mutagenic activity of airbone particulate materials collected at six different points in the industrial area of Ohmuta and the residential area Fukuoka. Tests were done in presence of rat-liver S-9 fraction isolated from rats that had been treated with Aroclor 1254. When the number of revertant colonies per plate was plotted against the amount of methanol extract of particulate air pollutants, using strain TA98, approximately linear relationships were observed for active samples. Generally, mutagenic activity of the samples increased in proportion to the density of air pollutants. In our system, 38--349 microng of methanol extract, from 0.225--4.51 m3 of air from the factory districts in Ohmuta City gave 100 his+ revertants per plate. On the other hand, 54--2300 microng of air pollutants, from 1.29--14.1 m3 of air from the residential districts in Fukuoka City, gave a comparable activity. Every sample from each area had mutagenic activity. Chemicals in air pollutants were fractionated by alumina column chromatography and identified by gas chromatography and mass spectrometry. More than 28 compounds, including 12 unknown substances were identified as polycyclic hydrocarbons. Twelve of these compounds are already known to be carcinogens and to induce reversions to histidine independence in strain TA98 of Salmonella.